BACKGROUND: Uncontrolled-rate freezing in -80 ℃ refrigerator is convenient, while controlled-rate freezing in -196 ℃ liquid nitrogen is reliable and long-term, the combination of the two can simplify the process and has been successfully used in clinics. OBJECTIVE: To explore the influences of different cryoprotectants by ladder-style freezing from -80 ℃ low temperature refrigerator to liquid nitrogen on the cryopreservation of hemopoietic stem cells. METHODS: The experiments were divided into four groups: 10% dimethyl sulfoxide (DMSO) group, 5% DMSO combined with 3% hydroxyethyl starch group, 5% DMSO combined with 0.25 mol/L trehalose group, 5% DMSO combined with 3% hydroxyethyl starch and 0.25 mol/L trehalose group. Peripheral hemopoietic stem cells were cryopreserved by ladder-style freezing from -80 ℃ low temperature refrigerator to liquid nitrogen. The ultrastructural changes were examined by transmission electron microscopy, the expressions of Annexin-V, PI and Caspase-3 were detected by flow cytometry. RESULTS AND CONCLUSION: There was no significant difference in survival rate, apoptotic rate and necrotic rates of the cryopreserved cells in the four groups (P > 0.05). The ultrastructural changes had no significant difference under the transmission electron microscopy. The viability was more than 90% in frozen-thawed mononuclear cell colonies, and the apoptosis was roughly 50% in the frozen-thawed CD45+ cell population, which contained many mature cells. Of hemopoietic stem cells, early stage cells have greater resistance to damage of cryopreservation than late stage cells. It is concluded that the addition of hydroxyethyl starch or trehalose into DMSO exhibits no synergistic protective effect on the cryopreservation of hemopoietic stem cells.

Toll-like receptor 5 (TLR5) is responsible for the recognition of bacterial flagellin in vertebrates. In the present study, the first TLR5 gene in duck was cloned. The open reading frame (ORF) of duck TLR5 (dTLR5) cDNA is 2580 bp and encodes a polypeptide of 859 amino acids. We also cloned partial sequences of myeloid differentiation factor 88, 2'-5'-oligoadenylate synthetase (OAS), and myxovirus resistance (Mx) genes from duck. dTLR5 mRNA was highly expressed in the bursa of Fabricius, spleen, trachea, lung, jejunum, rectum, and skin; moderately expressed in the muscular and glandular tissues, duodenum, ileum, caecum, and pancreas; and minimally expressed in the heart, liver, kidney, and muscle. DF-1 or HeLa cells transfected with DNA constructs encoding dTLR5 can activate NF-kappaB leading to the activation of interleukin-6 (IL-6) promoter. When we challenged ducks with a Herts33 Newcastle disease virus (NDV), mRNA transcription of the antiviral molecules Mx, Double stranded RNA activated protein kinase (PKR), and OAS was up-regulated in the liver, lung, and spleen 1 and 2 days post-inoculation.
2',5'-Oligoadenylate Synthetase/genetics/metabolism ; Animals ; Cell Line ; *Cloning, Molecular ; Ducks ; Gene Expression Regulation/*physiology ; Humans ; Immunity, Innate ; Myeloid Differentiation Factor 88/genetics/metabolism ; Myxovirus Resistance Proteins/genetics/metabolism ; Newcastle Disease/metabolism ; Newcastle disease virus/classification ; RNA, Messenger/genetics/metabolism ; Species Specificity ; Toll-Like Receptor 5/genetics/*metabolism

Objective To explore the role of TMAO from gut microbiota in non-alcoholic fatty liver disease(NAFLD),we detected the serum level of TMAO and its precursor metabolites in NAFLD,as well as the expression level of Eubacterium rectum,Bacteroidetes multiforme,Lactobacillus and bifidobacterium in the intestinal flora.Methods We collected 118 subjects and divided into NAFLD group(86 cases)and healthy control group(32 cases)randomly.We also detected the serum level of TMAO and its precursor metabolites in subjects by high performance liquid chromatography tandem mass spectrometry detection(LC-MS),and the expression of target bacterial DNA was detected by qRT-PCR.Results Serum TMAO,TMA and choline levels were significantly increased in NAFLD(P<0.05),and liver fat content was positively correlated with TMAO(P<0.05).The expression level of Lactobacillus and Eubacterium rectum in NAFLD group were increased(P<0.05);the expression level of Bifidobacterium and Bacteroides multiform were decreased(P<0.05).The serum TMAO level was positively correlated with Eubacterium rectum(r=0.280,P<0.05),and negatively correlated with Bifidobacterium(r=-0.332,P<0.05).Conclusion The level of TMAO in serum shows a positive correlation with NAFLD.The structure of intestinal flora in individuals with NAFLD is altered and linked to TMAO.This suggests that the intestinal flora may have a significant impact on the development of NAFLD through TMAO.

Objective:To explore the clinical and genetic characteristics of two children with a clinical diagnosis of Treacher Collins syndrome (TCS).Methods:Whole-exome sequencing was used to screen potential variants in the two children. Confirmation of suspected variants was performed through Sanger sequencing , multiplex ligation dependent probe amplification and real-time PCR in probands and their parents.Results:A heterozygous deletion variant, c. 4357_4360delGAAA, was detected in case one, while was de novo and verified by Sanger sequencing. Thevariant was classified as pathogenic(PVS1 + PM2+ PM6)according to ACMG guideline. The heterozygous deletion of exon 1-7 was seen in the same gene in case 2, which MLPA verified as heterozygous deletion of exon 1-6. This deletion was inherited from the father with a normal phenotype, and the father’s TCOF1 gene was suspected to be chimeric heterozygous deletion of exon 1-6 verified by MLPA. Conclusion:The identified variants in the TCOF1 gene probably underlie the two cases of TCS. There was no apparent correlation between genotype and phenotype. In addition, it shows a high interfamilial variability ranging from normal to full presentation of TCS. Genetic detection provided clinical diagnosis and genetic counselling for TCS patients .

To investigate the prevalence and epidemiological characteristics of diabetic retinopathy (DR) in Yunnan Province, explore its risk factors, and provide a basis for the prevention and treatment of chronic complications of diabetes mellitus (DM). This is a large cross-sectional study, in all, 1 524 DM patients in 16 communities and villages of Yunnan Province who were registered in health service centers were included in this study from August to November 2019. All patients completed a uniform questionnaire, anthropometric measurements, biochemical measurements, and auxiliary examinations. Logistic regression analysis was used to screen the risk factors of DR. The prevalence rates of DR, mild non-proliferative DR (mild-NPDR), and referable DR (RDR) were 16.0% (244/1 524), 4.5% (69/1 524), and 11.5% (175/1 524), respectively. Glycated hemoglobin A 1c (HbA 1c)≥7.0% was the risk factor of mild-NPDR ( OR=1.872, 95% CI 1.055-3.323) and RDR ( OR=4.821, 95% CI 2.917-7.969). Blood pressure≥130/80 mmHg (1 mmHg=0.133 kPa) was the risk factor of mild-NPDR ( OR=1.933, 95% CI 1.112-3.358) and RDR ( OR=1.505, 95% CI 1.063-2.130). In Yunnan Province, 16.0% DM patients had accompanying DR, wherein about 71.7% of them required an ophthalmology referral, and the high incidence of RDR in DM patients was associated with poor control of blood glucose and blood pressure.

Objective To explore the relationship between iodine and fluoride content in drinking water and the incidence of adult thyroid nodules in Cangzhou,Hebei.Methods According to the previous reports on iodine and fluoride levels in drinking water in Cangzhou,from November 2016 to January 2017,Cangzhou was divided into high iodine,low iodine,normal iodine and fluorine,low iodine and high fluorine,high iodine and high fluorine areas,and according to the different contents of iodine and fluorine in drinking water,high iodine and high fluorine area was further divided into high iodine and high fluorine 1 (iodine:743.30 μg/L,fluorine:4.27 mg/L),2 (iodine:119.31μg/L,fluorine:4.67 mg/L) and 3 (iodine:105.30 μg/L,fluorine:1.64 mg/L) subareas.Subjects who lived for 20 or more years and aged 30 or older,without serious disease and not taken iodized salt were selected.Palpation was used to examine the size,texture,mass,tenderness and mobility of the thyroid gland.The boundary,internal echo,blood flow and quantity of nodules were observed and recorded by color Doppler.Results The prevalence difference of thyroid nodules [36.8％ (629/1 710),32.8％ (636/1 938),25.1％ (427/1 700)] in high iodine,low iodine and normal iodine and fluorine areas was statistically significant (x2 =55.597,P ＜ 0.05).The prevalences of thyroid nodules in both high iodine and low iodine areas were higher than that of normal iodine and fluorine area (P＜ 0.016 7).The prevalence difference of thyroid nodules [43.3％ (749/1 730),39.8％ (712/1 790),34.9％ (623/1 785)] in high iodine and high fluorine 1,2 and 3 subareas was statistically significant(x2 =26.220,P ＜ 0.05).Compared with low iodine area,the prevalence of thyroid nodules [41.2％ (735/1 785)] in low iodine and high fluorine area was increased (x2 =6.288,P ＜ 0.05).Conclusions Both high iodine and low iodine can induce thyroid nodules.In water source areas with high iodine content,both high iodine and high fluorine are the factors inducing thyroid nodules.The prevalence of thyroid nodules in low iodine and high fluorine area is significantly higher than that of low iodine area.